Coformycin is a known unusual nucleoside discovered by H. Umezawa et al. and obtained from the culture broth of Nocardia interforma (ATCC No. 21072) and Streptomyces kaniharaensis SF-557 (ATCC No. 21070) which are the microorganisms producing the nucleoside antibiotics formycins A and B (see Japanese patent publication No. 12278/70 and the "Journal of Antibiotics" Ser. A. 17:, 96 (1964)). Coformycin may also be obtained from the culture broth of Streptomyces lavendulae MA25-A2. Coformycin has interesting biological and physiological properties such as an activity to inhibit remarkably the enzymatic deamination of formycin A and adenosine by adenosine deaminase (see the "Journal of Antibiotics" Ser. A, 20: 227, (1967)). Accordingly, coformycin is a very useful substance not only for the analysis of causes of diseases involving the metabolism of nucleic acid, but also for the chemotherapy of certain diseases, including tumors. Coformycin has the following chemical structure: ##STR1## (see the "Journal of the American Chemical Society" 96: 4327 (1974)) and may be synthesized chemically from 9-.beta.-D-ribofuranosylpurine (see Japanese Pat. No. 875,639 and the "Journal of the American Chemical Society" 96: 4326 (1974)).
The nucleosides which are related to coformycin and which are separated according to the process of this invention include isocoformycin, 2'-deoxycoformycin and formycins. Isocoformycin has the formula: ##STR2## and is an unusual nucleoside which is chemically synthesized by the present inventors with co-inventors and which remarkably inhibits the enzymatic activity of adenosine deaminase (see Japanese patent application No. 110219/76 filed on Sept. 14, 1976). 2'-Deoxycoformycin is an unusual nucleoside discovered by certain American researchers and isolated from the culture broth of Streptomyces antibioticus and is an inhibitor against the enzymatic activity of adenosine deaminase (see the "Journal of Heterocyclic Chemistry" 11, 641 (1974)).
2'-Deoxycoformycin has the following formula: ##STR3##
Hithertofore, the isolation and purification of coformycin from its related nucleosides usually required very troublesome operations either when coformycin was to be recovered from the culture broth of the Streptomyces strains, or when coformycin was to be recovered from the reaction mixture as formed in the chemical synthesis thereof. For instance, the culture broth of the coformycin-producing strains usually contains formycins A and B and other nucleosides in addition to the desired coformycin, and coformycin itself is very unstable under acidic conditions. Owing to these facts, it was normally required in the isolation and purification of coformycin to repeat chromatographic operations by adsorption and elution using a strong cation-exchange resin, activated carbon and a cation-exchanger such as a cross-linked dextran having basic function, for example, DEAE-Sephadex (a product of Pharmacia Co., Sweden) as described in Japanese patent publication No. 12278/70.
Accordingly, we have researched an attempt to devise an improved process for the isolation and purification of coformycin in order to recover coformycin in isolated and pure state from an aqueous solution of a crude powder comprising coformycin together with at least one of its related nucleosides. As a result, we have now found that coformycin can be isolated and purified in a facile way and in a high yield when a column chromatographic process is applied to said aqueous solution using a cation-exchanger having partially activated carboxylic groups as the ion-exchange function, and that coformycin can then be obtained easily in a pure crystalline form. It is to be noted that any cation-exchanger having carboxylic groups as the ion-exchange function has hithertofore never been employed for the chromatographic isolation and purification of coformycin from its related nucleosides. Furthermore, we have found that the chromatographic process using the cation-exchanger having partially activated carboxylic groups is generally applicable to said aqueous solution containing at least two of coformycin and its related nucleosides in order to separate them from each other.